Alginates are well known as versatile materials for cell encapsulation because of their ability to form highly biocompatible and strong gels under physiologic conditions at constant temperature (Skjåk-Bræk G, and T. Espevik Carbohydrates in Europe 1996; 14: 19-25; and Strand B L, et al. Minerva Biotecnologica 2000; 12: 223-233, which are incorporated herein be reference). Transplantation of alginate based tissue constructs may be useful in the treatment of a large number of diseases. Alginates can be used to entrap cells within microbeads, thus protecting the cells against immune attack from the host and physical stress. (Skjåk-Bræk G, and T. Espevik supra; Strand B L, et al supra; Yang H, et al. Cell Encapsulation Technology and Therapeutics. Boston, Birkhäuser, 1999: pp. 3-17; Uludag H, et al. Adv Drug Deliv Rev 2000; 42: 29-64; Orive G, et al. Nature Medicine 2003; 9: 104-107; Emerich D F and H C Salzberg Cell Transplant 2001; 10: 3-24; Sambanis A, Diabetes Technol Ther 2000; 2: 81-89 and Lanza R P, and D K Cooper. Molecular Medicine Today 1999; 4: 39-45, which are incorporated herein be reference.) Cells entrapped within alginate beads excreting therapeutic molecules may be used as implantable bioreactors in the treatment of a large variety of diseases, including cancer, diabetes, Parkinson's disease, chronic pain and liver failure (Emerich D F and H C Salzberg supra; Lanza R P, and D K Cooper supra; Wang T, et al. Nat Biotechnol 1997; 15: 358-362; Read T-A, et al. Nat Biotechnol 2001; 19: 29-34; Cai J, et al. Hepatology 2002; 36: 386-394; Canaple L, et al. Ann NY Acad Sci 2001; 944: 350-361; Glicklis R, et al. In: Ikada Y, Okano T (Eds.). Tissue Engineering for Therapeutic Use. 1999: pp. 119-131; Emerich D F. Cell Transplant 2002; 11: 1-3; Emerich D F, et al. Neurosci Biobehav Rev 1992; 16: 437-447; Hagihara Y, et al. Cell Transplant 1997; 6: 527-530; Risbud M V and R R Bhonde J Biomater Sci Polymer Edn 2001; 12: 1243-1252, which are each incorporated herein by reference.) Therefore, alginates are now widely used as immobilizing materials for cells or tissue in the development of bioreactor systems for therapeutic use.
Alginates are also being studied as a biostructure materials in other types of medical applications. Dependent of the manufacturing process alginates may take various forms such as pastes, sponges, fibers, rods and tubes. Alginate sponges are being studied as materials for cell transplantation (Miralles G, et al. Journal of Biomedical Materials Research 2001; 57: 268-278; and Shapiro L and S. Cohen Biomaterials 1997; 18: 583-590, which are each incorporated herein by reference.) and nerve regeneration (Sufan W, et al. Journal of Neurotrauma 2001; 18: 329-338; Kataoka K, et al. J Biomed Mater Res 2001; 54: 373-384; and Hashimoto T, et al. Exp Brain Res 2002; 146: 356-368, which are each incorporated herein by reference.) Furthermore, alginate pastes containing chondrocytes have been injected into children as successful treatment of urethral reflux problems (Diamond D A and A A Caldamone J Urol 1999; 162: 1185-1188, which is incorporated herein by reference) and implants of chondrocytes in alginate gelled in situ by the addition of gelling solution directly into cartilage defects are promising (Fragonas E, et al. Biomaterials 2000; 21: 795-801, which is incorporated herein by reference.)
For applications involving cells in direct contact with the alginate structure, interactions between the cells and the alginate matrix may be crucial. In addition with respect to alginate bioreactor systems in particular, a major obstacle may be the selection and availability of sources of producer cells. As an alternative to processing fresh organs shortly prior to medical use, there are some advantages to growth of cells in vitro as an unlimited source for bioreactor production. Such cells can be genetically manipulated for better properties and the ability to produce therapeutic products.
Generally, for alginate entrapped proliferating cells, cell growth must in some way be controlled. It has been established that cell growth within the alginate gel matrix is dependent of the type of gel network (Constantinidis I, et al. Biomaterials 1999; 20: 2019-2027; and Stabler C, et al. Biomaterials 2001; 22: 1301-1310, which are each incorporated herein by reference), but the growth is also cell type dependent (Rokstad A M, et al. Cell Transplant 2002; 11: 313-324, which is incorporated herein by reference). Cells entrapped in weaker alginate gels containing a low content of guluronic acid have been shown to grow more rapidly as compared to cells entrapped in stronger i.e. high guluronic acid content gels (Constantinidis I, et al. supra; and Stabler C, et al. supra). As a result of cell growth and formation of colonies within the gel network, beads may disrupt and leakage of cells from the beads may occur (Constantinidis I, et al. supra; and Stabler CL, et al Ann N Y Acad Sci 2002; 961:130-133, which is incorporated herein by reference). Thus, a particular problem when using proliferating cells in alginate beads is that the cells continue to grow and proliferate, and the bead disruption and cell leakage that occurs exposes the cells to the immune system.
Animal cells are highly specialized in responding to and interacting with adjacent cells and extracellular matrixes. Such responses are controlled by specific genes. It has been demonstrated that collagen, a major normal extracellular matrix component, inhibits cells from entering into apoptosis and thereby provide a substrate for cell survival and differentiation (O'Connor S M, et al. Neurosci Lett 2001; 304: 189-193, which is incorporated herein by reference). The molecular mechanisms behind the interaction between cells and an alginate matrix are, however, unknown. While entrapped cells may form spheroid-like colonies within the gel network, it has also recently been demonstrated that cells may grow attached to alginate gel surfaces (Wang L, et al. Biomaterials 2003; 24: 3475-3481, which is incorporated herein by reference). It was established that rat bone marrow cells may grow on alginate gel surfaces in vitro without any chemical modification of the gel substrate (Wang L, et al. supra). In contrast to what was previously observed for cell growth within the alginate matrix, Wang et al also found a higher proliferation rate on gels of alginate with a high as compared to low content of guluronic acid. However, the lack of ability for C2C12 myoblasts to grow on non-chemically modified alginate surfaces has also been observed, while RGD peptide sequences bound to the alginate substrate allowed cell growth (Rowley J A et al. Journal of Biomedical Materials Research 2003; 60: 217-223; and Rowley J A, et al. Biomaterials 1999; 20: 45-53, which are incorporated herein by reference). These workers, however, also found best proliferation of myoblasts on alginate gels made with alginates containing a high content of guluronic acid.
Another common problem in applications involving implantation of alginate beads into in to animals is the growth of fibroblast and macrophages at the surface of selected beads (Vandenbossche G M R, et al. J Pharm Pharmacol 1993; 45: 115-120; Rokstad A M, et al. Ann N Y Acad Sci 2001; 944: 216-225; and Siebers U, et al. Journal of Molecular Medicine 1999; 77: 215-218, which are each incorporated herein by reference). This problem also occurs when other foreign bodies, such as devices, are implanted. Better knowledge about cell growth and attachment behavior in contact with the alginate matrix is therefore clearly needed.
There is a need to provide compositions comprising proliferating cells encapsulated in alginate and methods of using such compositions in which the growth and proliferation of the cells is controlled, thereby preventing bead disruption, cell leakage and the immune response that follows.
There is a need for compositions comprising cells encapsulated in alginate and methods of using such compositions wherein the cells excrete therapeutic molecules.
There is a need for compositions comprising cells encapsulated in alginate and methods of using such compositions in the treatment of diseases.
There is a need to provide implantable compositions and devices and methods of using such implantable compositions and devices in which the growth of unwanted host cells on the surface of such implantable compositions and devices is controlled.